The long-term goal of the proposed project is to delineate the molecular genetics of infantile neuronal ceroid lipofuscinosis (INCL) and to understand how deficiency in a key lysosomal enzyme, palmitoyl-protein thioesterase, causes the neurodegenerative phenotype. INCL is a devastating disorder characterized by normal development to the age of one year, followed by relatively rapid onset of blindness, seizures, and loss of cognitive and motor function. The EEG is isoelectric by age 3. Cortical areas of the brain are disproportionately affected, so that these children typically persist in a vegetative state for a decade or longer. In contrast to other lysosomal storage diseases that affect the brain, severe neuronal loss rather than neuronal storage dominates the pathology. INCL is caused by mutations in the gene encoding palmitoyl-protein thioesterase (PPT1), a lysosomal enzyme that was purified and cloned in the course of the study of the turnover of lipids on fatty acylated proteins. Lipid thioesters derived from acylated proteins accumulate in cells derived from INCL patients. Therefore, it is hypothesized that these lipid thioesterase are neurotoxic and their accumulation is central to disease pathogenesis. A second lysosomal thioesterase (PPT2) exists in the brain but apparently cannot compensate for the deficiency in PPT1.
Four specific aims are proposed. First, the molecular basis for INCL phenotypes will be explored through the characterization of recombinant mutant PPT enzymes. Particular attention will be paid to late-onset phenotypes that were uncovered during the previous funding period. Second, a mouse model of INCL lacking PPT1 will be created and characterized and the storage material accumulating in the brains of these mice will be analyzed. Third, the function of PPT2 will be explored through the created and characterization of a PPT2-deficient mouse. Fourth, mice lacking both PPT1 and PPT2 will be created and characterized. The results from these studies will lead to unique new insights into lysosomal thioester metabolism, neurodegeneration, and aging.
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